Genetic alterations of KIT during clonal expansion and subsequent acquisition of resistance to toceranib in a canine mast cell tumor cell line

J Vet Pharmacol Ther. 2019 Nov;42(6):673-681. doi: 10.1111/jvp.12816. Epub 2019 Sep 25.

Abstract

One of the potential mechanisms underlying acquired resistance to toceranib in canine mast cell tumor (MCT) is the emergence of a secondary mutation in the KIT gene. Here, genetic alterations of KIT during clonal expansion and subsequent acquisition of resistance to toceranib were investigated in the toceranib-susceptible canine MCT cell line VI-MC, which carries a KIT-activating mutation resulting in a predicted p.(Asn508Ile) amino acid change in the receptor tyrosine kinase protein KIT. Two sublines were cloned from VI-MC and toceranib-resistant sublines then were established by continuous exposure to toceranib. The mutation status of KIT in parental VI-MC and its sublines was investigated using next-generation sequencing (NGS). Additionally, effects of secondary mutations on toceranib sensitivity in p.(Asn508Ile)-mutant KIT were examined. KIT secondary mutations, including those encoding p.(Asn679Lys)-, p.(Asp819Val)-, and p.(Asp819Gly)-mutant KIT, that confer toceranib insensitivity to p.(Asn508Ile)-mutant KIT emerged only in toceranib-resistant VI-MCs. These mutations were not detected by NGS in the parental VI-MC line or in the toceranib-naive cloned VI-MCs, although the parental line and sublines exhibited genetic heterogeneity in KIT that may have been caused by genetic evolution during clonal expansion. VI-MC clones with these secondary mutations in KIT appear to have arisen from subclones during treatment with toceranib rather than being pre-existing. However, further study using a higher resolution technique will be needed to confirm the developmental mechanism of KIT secondary mutation in canine MCT cells with acquired resistance to toceranib.

Keywords: KIT; dog; genetic alterations; mast cell tumor; resistance; toceranib.

MeSH terms

  • Animals
  • Antineoplastic Agents / pharmacology
  • Cell Line, Tumor
  • Cloning, Molecular
  • Dogs
  • Drug Resistance, Neoplasm / genetics*
  • Gene Expression Regulation, Neoplastic / drug effects
  • HEK293 Cells
  • Humans
  • Indoles / pharmacology*
  • Proto-Oncogene Proteins c-kit / genetics*
  • Pyrroles / pharmacology*

Substances

  • Antineoplastic Agents
  • Indoles
  • Pyrroles
  • toceranib phosphate
  • Proto-Oncogene Proteins c-kit